The ability of an organism to withstand bacterial invasion depends upon sensitive and specific molecular systems. The molecules involved in these systems are designed to recognize specific bacterial products and trigger rapid responses to small numbers of invading bacteria.
Innate recognition systems include highly conserved “pattern recognition” host molecules that detect and respond to highly conserved and structurally unique microbial molecules. The best-studied example of such an innate system is the machinery engaged in recognition of endotoxins, which are unique surface glycolipids of Gram-negative bacteria.
Potent pro-inflammatory cellular responses to endotoxin are mediated through activation of a member of the Toll-like receptor family of proteins, Toll-like receptor 4 (TLR4; Beutler et al., 2003; Means et al., 2000; and Ulevitch et al., 1999). An important feature of TLR4-dependent cell activation by endotoxin is its extraordinary sensitivity, permitting timely host responses to small numbers of invading Gram-negative bacteria, essential for efficient host defense (Beutler et al., 2003; Means et al., 2000; and Ulevitch et al., 1999).
TLR4 contains a leucine-rich extracellular domain involved in ligand recognition, a transmembrane region, and an intracellular domain responsible for triggering signalling pathways that result in activation of genes of the innate immune defense system (Beutler et al., 2001; and Medzhitov et al., 1998). Maximal potency of TLR4-dependent cell activation by endotoxin requires four different extracellular and cell surface host proteins: lipopolysaccharide (LPS) binding protein (LBP), CD14, MD-2 and TLR4 (Beutler et al., 2003; Miyake et al., 2003; and Ulevitch, 2000).
TLR4 requires MD-2 for CD14-dependent cellular response to low concentrations of endotoxin, but neither the precise nature of the ligand that binds to TLR4 nor the role of MD-2 has been fully defined. MD-2, either endogenously expressed or exogenously added, associates with TLR4 on the cell surface (Viriyakosol et al., 2001; Schromm et al., 2001; Visintin et al., 2001; Re et al., 2002; Akashi et al., 2003; Visintin et al., 2003; and Re et al., 2003) and its endogenous expression is needed for optimal surface expression of TLR4. TLR4 responsiveness to endotoxin is disrupted by point mutations of MD-2 (Schromm et al., 2001; Kawasaki et al., 2003; Ohnishi et al., 2001; and Mullen et al., 2003) (e.g., C95Y, Lys128 and Lys132) despite surface expression of TLR4/MD-2 complexes.